The broad long-term objective of this proposal is to examine the source sphingosine-1-phosphate (S1P) in the pathology involved in inflammatory bowel disease (IBD), and target its synthetic enzyme, sphingosine kinase 1 (SKI) with specific inhibitors as a potential novel therapeutic approach to the disease. In Specific Aim 1, we will establish that colon tissue SK1 is the source of SI P in IBD. This will be done by using a mouse bone marrow chimera model in conjunction with a mouse model of ulcerative colitis. We will (i) establish bone marrow chimeric mice, by irradiating both wild-type and sphingosine kinase-1 knockout mice and reconstitute their blood with either host blood or blood from the opposite strain. We will then (ii) administer DSS to induce colitis in bone marrow chimeric mice, and examine parameters of disease: such as weight loss, colon shortening, splenomegaly, histologic markers of disease, blood and tissue SK1 activity, as well as sphingolipid changes. Next we will (Hi) examine the cellular inflammatory responses with flow cytometry and the underlying mechanisms of inflammation in colon tissue, such as TNFor and C0X2 production from DSS-administered, bone marrow chimeric mice. These studies have an obvious link to human health and should firmly establish the source of SI P in IBD to allow potential targeting of therapy. In Specific Aim 2 we will develop novel SKI inhibitors as potential new therapeutic modalities for the treatment of IBD. This aim will test the sub-hypothesis that inhibiting SKI will ameliorate the severity of IBD. To this end we will (1) evaluate our newly developed SK1 inhibitors as to their ability to inhibit TNFa activation of SK1 and downstream signaling in FHC human colon epithelial cells. We will examine these downstream effects by Real Time-RTPCR and western blotting. We will also (ii) treat mice with SKI inhibitors by either i.p. or i.v. injection and evaluate tissue and blood SKI as well as their response to DSS- induced colitis as done for Specific Aim 1. These studies advance SKI inhibitors as novel therapeutic approaches to IBD. PUBLIC HEALTH RELEVANCE: This research is directly related to human health and disease. Current therapies for IBD involve general immunosuppression and the use of steriods, causing undesirable side effects in patients leading to issues of compliance. More specific targets with less side effects are needed for treatment of this disease. This research is targeted at finding just that, a novel therapeutic that inhibits the cellular mechanism of disease and improving patient health.